Cytokinin has various types of physiological activities such as promotion of cell division, regulation of cell cycle, retardation of senescence, or activation of axillary buds outgrowth, and thus it is an extremely important plant hormone for controlling the quantitative productivity of crops.
As a basic structure, cytokinin has a prenyl group containing 5 carbon atoms at the nitrogen atom on the position 6 of adenine. Depending on the difference of a side chain structure, cytokinin includes trans-zeatin (tZ), isopentenyladenine (iP), etc. Based on the findings from conventional studies regarding a cytokinin-metabolizing system, it has been considered that the synthesis of active cytokinin (a base form) is carried out via at least the following 3 steps of reactions. First, in a first reaction of cytokinin synthesis, nucleotide cytokinin is produced as a result of a condensation reaction between adenine nucleotide and dimethylallyl diphosphate (DMAPP). This nucleotide form does not have activity as cytokinin. However, in a second reaction, such nucleotide form is converted to nucleoside cytokinin by the action of dephosphorylating enzyme. Thereafter, ribose is dissociated by the action of nucleosidase, and thus it is converted to an active cytokinin molecule of a base form (Non-Patent Document 1; Review: Sakakibara, H. (2006) Cytokinins: Activity, biosynthesis and translocation. Annu. Rev. Plant Biol. 57: 431-449.). As a matter of fact, in a study at an enzyme level in 1980s, enzymes catalyzing each of the aforementioned reactions have been suggested. An enzyme gene (IPT) catalyzing the first reaction (condensation reaction), namely, the synthesis reaction of nucleotide cytokinin, was identified in 2001 (Patent Document 1: WO2002/072818). However, enzyme genes catalyzing the second reaction (activation reaction) consisting of the remaining 2 steps have not yet been identified. Even if the entities of such genes are identified, in order to artificially regulate the amount of cytokinin as a base form generated, two steps, namely, a step of converting a nucleotide form to a nucleoside form and a step of converting the nucleoside form to a base form, must be simultaneously modified. Thus, it is necessary to identify enzyme genes catalyzing the two reactions and then to simultaneously regulates the identified enzyme genes. Such operation is not easy. On the other hand, it is also possible to regulate the amount of active cytokinins by regulating an enzyme gene (CKX) catalyzing a cytokinin-degrading reaction (Non-Patent Document 2: Werner T, Motyka V, Strnad M, Schmulling T. (2001) Regulation of plant growth by cytokinin. Proc Natl Acad Sci USA 98: 10487-10492; Non-Patent Document 3: Ashikari, M., Sakakibara, H., Lin, S.-Y., Yamamoto, T., Takashi, T., Nishimura, A., Angeles, E. R., Qian, Q., Kitano, H. and Matsuoka, M. (2005) Cytokinin oxidase regulates rice grain production, Science, 309: 741-745.). However, a technique of directly regulating the amount of active cytokinin synthesized has not yet been developed so far. In addition, the amount of a base form is increased by excessive expression of IPT, but such a method is problematic in that the total amount of cytokinin is extremely increased due to such excessive expression of IPT and the form as a plant body is also significantly changed (Non-Patent Document 4: Zubko E, Adams C J, Machaekova I, Malbeck J, Scollan C, Meyer P. 2002. Activation tagging identifies a gene from Petunia hybrida responsible for the production of active cytokinins in plants. Plant J. 29: 797-808.).